This document discusses microplate assays, which are a key platform for high-throughput screening in biomedical research and pharmacology. Microplates were developed in the 1990s and contain rows and columns of wells in formats of 6, 24, 96, 384, or 1536 wells. They are used for enzyme-linked immunosorbent assays and high-throughput screening of large chemical libraries to identify potential drug candidates. Microplate readers are used to analyze the assays automatedly and efficiently.

1. M I C R O P L AT E A S S AY S : T H E K E Y
P L AT F O R M F O R H I G H T H R O U G H P U T
SCREENING IN BIOMEDICAL
RESEARCH AND PHARMACOLOGICAL

2. Microtiter Plate
•
develop in 1990s
•
2:3 rectangle matrix with wells
•
6, 24, 96, 384, 1536 wells
•
used for Enzyme-linked Immunosorbent Assay, High
Throughput Screening, storage
•
Flat-shaped, U-shaped, V-shaped

3. Well shape

4. ELISA

5. Microplate Reader
taken from Synergy

6. Reusing plate
taken from © 2014 Kinesis Ltd.
taken from Labotal Scientific Equipment (1997) LTD

7. Materials
•
polyprolyene- for wide change in temperature
environment
•
polycrabonate- disposable,
•
Cyclo-olefins

8. High Throughput Screening
an optimized, miniaturized assay format that enables the
testing of > 100,000 chemically diverse compounds per
day.

9. Biotechnology Research Institute
The Hong Kong University of Science and Technology
•
TCMs, including those for treatment of Alzheimer’s
disease, neuro-protection, stroke, and sleep disorders

10. taken from RNAi @ NIH Chemical Genomics Center

11. History of Developing a Drug
!
Drug discovery
!
Initial characterisation
!
Pre-clinical trials
!
Regulatory approval sought to start
trials in humans
!
Clinical trials Phases I, II, III
!
Submission of marketing/manufacturing
authorization application to regulatory authorities
!
Regulatory authorities review
information and grant (or refuse) licences
!
Product goes on sale
!
Post-marketing surveillance
!
Library of compounds
!
In vitro screening: human/animal
receptor/enzyme assay; reporter system
!
Hits/lead
!
Biochemical, tissue or animal model of
function
!
lead
!
Animal model of therapeutic target
!
ADME, formulation, acute toxicology

12. High throughput screening for drug discovery
high throughput screening (HTS) is number one tool for early-stage drug discovery
!
HTS is process by which large numbers of compounds are rapidly tested for their
ability to modify the properties of a selected biological target.
Goal is to identify ‘hits’ or ‘leads’
- affect target in desired manner
- active at fairly low concs (∴ more likely to show specificity)
- new structure
!
!
HTS = 50,000-100,000 cpds screenings per day!!!

13. taken from Pete the Blogger

14. !
Aim of screening is to find progressible hits, not to discover
the lead molecule itself
!
!
The majority of drug targets are
a) G-protein coupled 7 TM receptors (est total 5000)
b) nuclear receptors
c) ion channels
d) enzymes
(est total >150)
(est total 1000)
(est total uncertain)

15. Assays
Definition - a test system in which biological activity can be detected
•
Aequorin assays
•
Colorimetric assays
•
Membrane potential assays
•
Intracellular calcium flux assays
•
Fluorometric assays
•
Luciferase reporter gene assays
•
Fluorescent and radiolabeled ligand binding assays

16. Fluorescent Polarization
Assay
taken from GlycoForum

17. Aequorin assay

18. Luciferase Reporter Gene Assay
from Thermo Scientific

19. Virtual Screening
•
computational technique used in drug discovery to search
libraries of small molecules in order to identify those structures
which are most likely to bind to a drug target, typically a protein
receptor or enzyme

20. •
Ligand based- logic based
rules feature substructures
and chemical properties of
the candidate ligands
•
Structure based- virtually
docking candidate ligands to
the receptors

21. The End